It has become increasingly evident that the thermal response of tumor cells in vivo is considerably different from that of cells in vitro probably because tumor cells in vivo are chronically exposed to acidic, hypoxic or nutritionally deprived intratumor suboptimal conditions, which complicates the use of in vitro information in heating tumors in vivo in the clinic. The overall objective of this proposal is to determine the relative importance of environmental factors, such as pH, pO2 and glucose supply, on the thermal response of tumor cells in vivo. Since the microenvironment of tumors in vivo cannot be precisely controlled, tumors will be dissociated into single cells or tumor fragments, and the in vivo-derived cells and fragments will be subjected to a controlled environment in vitro, in which the thermal characteristics of in vivo cells are preserved even following the dispersion. Four specific aims are planned: (1) Heat response of in vivo-derived tumor cells will be studied under various environmental conditions in vitro and the results will be compared with that of cells adapted to the culture conditions in vitro. Since the pH effect was already studied, the effects of glucose and hypoxia with or without combination of low pH will be studied. (2) The mechanisms involved in the transition of heat sensitivity of in vivo-derived cells to a resistant state will be studied in detail, using metabolic inhibitors which are active on H+-ion distribution, glutathione synthesis and energy production in cells. (3) Concomitant determination of the biochemical effect of the specific inhibitors (Specific Aim 2) will also be carried out. Cellular glutathione contents and energy charge will be determined by using HPLC. (4) In order to correlate the above information (Specific Aims 1-3) with the situations of tumors in vivo, the condition of the intratumor milieu will be examined. The pH and pO2 distributions in tumors either prior to, during, or after heating will be determined using glass microelectrodes and tumor blood flow will be measured using a laser Doppler flowmeter. The above proposed studies will be conducted with two different mouse tumor models, SCK mammary carcinoma and RIF-1 fibrosarcoma in which the vascular response of the two tumors are quite different from each other. Such information will contribute to a better understanding of factors influencing tumor response to heat as an organized tissue and for developing treatment protocols for the more effective use of hyperthermia in the clinic.